A variety of methods have been developed for determining substances capable of being bound specifically, generally immunochemical substances, including antigens, antibodies, haptens and other low molecular weight substances. In one approach, binding reagents capable of binding with a target analyte are bound to an insoluble support. When the support is contacted with the sample containing the analyte, the analyte is bound to the binding reagent, and is removed from the sample when the insoluble support is separated from the sample. Methods using insolubilizing supports include radioimmunoassay techniques, fluoroimmunoassays, enzyme immunoassays, transistor bridge probes, indium reflective surface methods, ultrasonic probes, and the like.
This approach is particularly useful in sandwich and competition immunoassays, wherein the insoluble support bearing the binding reagent conjugated with the analyte is further reacted or conjugated with labeled reagents which bind specifically with the analyte or unconjugated binding reagent. Measurement of the label bound to the insoluble support can be correlated with the amount of analyte in the sample. Binding of the labeled reagent exclusively with the analyte is intended in these procedures. However, a certain proportion of the labeled reagent inevitably binds directly with the insoluble support in a non-specific binding process. This "non-specific" binding introduces an error variable which contributes a background signal limiting the sensitivity of the assay method. This type of limit has become increasingly important with the introduction of labels yielding a high signal over the environmental background and the development of assays for low trace quantities of analytes.
As a partial solution to the problem of non-specific binding, an insoluble support to which a binding reagent has been bonded is sometimes treated with a solution containing a non-immune protein or amino acid polymer which coats the support surface, presenting a barrier to further protein binding by physical adsorption. This non-specific binding inhibitor reduces the non-specific binding, but with the insoluble supports previously used in heterogeneous sandwich and competition immunoassays, a substantial level of non-specific binding remained.
This invention is based on the discovery that polysilicon surfaces are substantially superior to surfaces previously used for sandwich and competition immunoassays because the non-specific binding of secondary reagents to the surface can be almost entirely inhibited with a coating of a suitable protein.
A wide variety of insoluble materials have been used or suggested for use as insoluble supports. Organic and inorganic polymers, both natural and synthetic, have been used. Examples of such polymers include polyethylene, polypropylene, polybutylene, poly(4-methylbutylene), butyl rubber, silastic polymers, polyesters, polyamides, cellulose and cellulose derivatives (such as cellulose acetate, nitrocellulose and the like), acrylates, methacrylates, vinyl polymers (such as polyvinyl acetate, polyvinyl chloride, polyvinylidene chloride, polyvinyl fluoride, and the like), polystyrene and styrene graft copolymers, rayon, nylon, polyvinylbutyrate, polyformaldehyde, etc. Other materials which have been used as insoluble supports are latices of the above polymers, silica gel, glass, paper, insoluble protein, metals, metalloids, metal oxides, magnetic materials, cermets and the like.